Abstract
Neutral proteases have broad application as additives in modern laundry detergents and therefore, thermostability is an integral parameter for effective production of protein crystals. To improve thermostability, the contribution of individual residues of Bacillus cereus neutral protease was examined by site-directed mutagenesis. The Lys11Arg and Lys211Arg mutants clearly possessed improved thermostabilities (Tm were 63 and 61 °C respectively) compared to the wild-type (Tm was 60 °C). MD simulations further revealed that the mutants had low RMSD and RMSF values compared to wild-type BCN indicating increased stability of the protein structure. Lys11Arg mutant particularly possessed the lowest RMSD values due to increased residue interactions, which resulted in enhanced thermostability. The mutants also displayed strong stability to most inhibitors, organic solvents and surfactants after incubation for 1 h. This study demonstrated Lys–Arg mutation enhanced thermostability of BCN and thus provides insight for engineering stabilizing mutations with improved thermostability for related proteins.
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Acknowledgements
This work was supported by the National Natural Science Foundation of China (Grant Nos. 21778024, 31870066, 31570085), the National Key Research and Development Program of China (Grant No. 2018YFA090039), National First-Class Discipline Program of Light Industry Technology and Engineering (Grant No. LITE 2018-06), Fundamental Research Funds for the Central Universities (Grant No. JUSRP51708A), the 111 Project (Grant No. 111-2-06), and the Priority Academic Program Development of Jiangsu Higher Education Institution. We further acknowledge the Collaborative guidance from lab mates and friends.
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Osire, T., Yang, T., Xu, M. et al. Lys–Arg mutation improved the thermostability of Bacillus cereus neutral protease through increased residue interactions. World J Microbiol Biotechnol 35, 173 (2019). https://doi.org/10.1007/s11274-019-2751-5
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DOI: https://doi.org/10.1007/s11274-019-2751-5